scholarly journals Development of an Albumin-Based PSMA Probe With Prolonged Half-Life

2020 ◽  
Vol 7 ◽  
Author(s):  
Teli Liu ◽  
Chen Liu ◽  
Yanan Ren ◽  
Xiaoyi Guo ◽  
Jinquan Jiang ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Half Life ◽  
Radiochemical Purity ◽  
Specific Activity ◽  
Tumor Burden ◽  
Tumor Bearing ◽  
Specific Membrane ◽  
Psma Inhibitor

Prostate-specific membrane antigen (PSMA) is an attractive target for the diagnosis and therapy of prostate cancer as it is specifically overexpressed in prostate cancer cells. Improving the circulation of radioligands in the blood is considered as an effective strategy that can improve tumor burden, which benefits detection of small lesions and improves the effect of PSMA radioligand therapy (PRLT). In this study, we introduced maleimidopropionic acid (MPA) to a PSMA-targeted tracer and developed Al18F-PSMA-CM, which targets human serum albumin (HSA) binding and PSMA. Al18F-PSMA-CM is evaluated in vitro and in vivo for stability, PSMA specificity, and biodistribution in 22Rv1 tumor-bearing mice. Al18F-PSMA-CM was prepared with a radiochemical purity of >99% and specific activity of 11.22–18.70 MBq/nmol. Al18F-PSMA-CM was stable in vitro and in vivo and prolonged circulation in blood with a binding ratio of 47 ± 3.2% and Kd value of 3.08 ± 0.45 nM to HSA. The uptake of Al18F-PSMA-CM in PSMA(+) 22Rv1 cells was increased in 2 h, and the uptake was blocked by a PSMA inhibitor, ZJ-43. The Kd value of Al18F-PSMA-CM to PSMA was 8.46 ± 0.24 nM. Al18F-PSMA-CM was accumulated in kidneys and 22Rv1 tumors [74.76 ± 15.42 and 6.16 ± 0.74 ID%/g at 2 h post injection (p.i.)], which were decreased by −80.0 and −84.3% when co-injected with ZJ-43. Al18F-PSMA-CM showed high PSMA specificity and accumulated in 22Rv1 tumors with increasing uptake in 4 h. MPA moiety showed the ability to prolong the half-life of tracers, and the MPA-conjugated tracer showed the potential to improve tumor uptake. MPA may be a choice to develop radiopharmaceuticals for PRLT of prostate cancer.

scholarly journals In Vitro Characterization of Rabbit Thrombin, Antithrombin III, and Thrombin-Antithrombin III Complex, and Determination of Their Survival Times in Vivo

1977 ◽  
Author(s):  
Christine N. Vogel ◽  
Kingdon S. Henry ◽  
Roger L. Lundblad
Keyword(s):  
Gel Electrophoresis ◽  
Half Life ◽  
Antithrombin Iii ◽  
Specific Activity ◽  
Sodium Dodecyl ◽  
In Vivo Studies ◽  
Survival Times ◽  
At Iii

Our intention is to study the interaction of rabbit thrombin with antithrombin III (AT-III) in vitro and in vivo. After activation of crude prothrombin with tissue thromboplastin and CaCl2, thrombin was purified and showed two species of thrombin with molecular weights of 36,000 and 39,000 daltons as determined by sodium dodecyl sulfate discontinuous gel electrophoresis. Rabbit AT-III was purified using a heparin agarose column and had a molecular weight of 55,000 daltons. The inhibition of thrombin by AT-III was followed by fibrinogen clotting assays and an AT-III-thrombin complex was observed on gel electrophoresis. For the in vivo studies both thrombin and AT-III were radiolabelled with Na125i using the solid state lactoperoxidase method and retained 99% of the pre-iodinated specific activity. Radiolabelled thrombin and a radiolabelled AT-III-thrombin complex were injected into different rabbits. The rate of removal of both was very similar with a half-life of approximately 9 hours. When radiolabelled AT-III was injected, the half-life was approximately 60 hours. Since the disappearance rate of thrombin more closely approximates that of the preformed AT-III-thrombin complex and is clearly shorter than the turnover rate of AT-III, the possibility is raised that thrombin combines in vivo with a native inhibitor such as AT-III and may in fact be removed from the circulation as a complex rather than as a native molecule.

In Vitro and In Vivo Characterization of Full-Length rFVIII Modified with PEG Via Coupling to Primary Amino Groups.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3147-3147 ◽  
Author(s):  
Peter L. Turecek ◽  
Jürgen Siekmann ◽  
Herbert Gritsch ◽  
Katalin Váradi ◽  
Rafi-Uddin Ahmad ◽  
...  
Keyword(s):  
Half Life ◽  
Hemophilia A ◽  
Specific Activity ◽  
Exchange Chromatography ◽  
Full Length ◽  
Thrombin Inhibitor ◽  
Knock Out ◽  
Knock Out Mice

Abstract Chemical modification of recombinant therapeutic proteins with PEG has been shown to enhance the biological half-life. Here we assess the effect of PEGylation on FVIII. Full-length rFVIII bulk drug substance from protein-free fermentation (Advate process, Baxter) was conditioned into a buffer suitable for coupling to polyethylene glycol succinimidyl succinate (linear PEG, 5 kDa PEG chain length). PEG was covalently bound by amine coupling preferentially to lysine residues of FVIII at neutral pH. PEG was removed by ion-exchange chromatography and the PEG-FVIII derivative was concentrated by ultra-diafiltration. The conjugates thus obtained retained about 30–40% of the activity of non-modified rFVIII. The specific activity decreased with the amount of PEG linked to the FVIII molecule. In SDS-PAGE and immunoblot studies PEGylated rFVIII showed a band pattern similar to unmodified FVIII with full-length, heavy chain fragments of 180 kDa and 120 kDa and the light chain fragment of 80 kDa. PEGylation also occurred to a high extent in the B domain of FVIII. All bands appeared broadened due to the attachment of polymeric PEG. The maintenance of functionality of FVIII was demonstrated by its potential to be activated and inactivated by thrombin. In the assay PEGylated and unmodified FVIII were incubated with 1 nM thrombin. Sub-samples were drawn at intervals up to 40 minutes and added to a mixture of FIXa, FX, phospholipid vesicles and Ca2+ containing a thrombin inhibitor. After 3 minutes incubation at 37°C the amount of activated FX (FXa) was measured using a FXa-specific chromogenic substrate. Unmodified rFVIII showed a typical picture of an immediate increase in FXa activity and a subsequent decline with no further FXa generation after 15 minutes. PEGylated rFVIII was activated to the same extent as unmodified FVIII but the decay in FXa generation was slower and did not reach the zero level, even 40 minutes after incubation. The formation of the typical thrombin cleavage fragments, with unmodified as well as PEGylated rFVIII, was demonstrated in a Western blot analysis. The slower inactivation by thrombin was also seen there. The pharmacokinetic properties of PEGylated rFVIII compared with rFVIII were investigated in hemophilia A knock-out mice. Both preparations were applied at a dose of 200 IU rFVIII/kg and groups of mice (n=5) were exsanguinated at several time points up to 24 hours. Terminal half-life for PEGylated rFVIII was calculated at 4.9 hours compared with 1.9 hours for unmodified rFVIII in hemophilia A knock-out mice. AUC was approximately doubled. These results indicate that rFVIII can be biochemically modified with PEG whilst at least partly retaining its major functions, but at the same time prolonging its survival in the circulation of hemophilic mice.

Preclinical evaluation of 99mtechnetium-HYNIC(tricine/TPPTS)-aca-bombesin(7–14) as a targeted imaging agent in prostate cancer.

2011 ◽  
Vol 29 (7_suppl) ◽  
pp. 122-122
Author(s):  
H. J. Ananias ◽  
Z. Yu ◽  
P. H. Elsinga ◽  
I. J. de Jong
Keyword(s):  
Prostate Cancer ◽  
Specific Activity ◽  
Human Prostate ◽  
Log P ◽  
P Value ◽  
Targeted Imaging ◽  
Human Prostate Cancer ◽  
Athymic Mice

122 Background: The peptide bombesin (BN) and derivates thereof show high binding affinity for the gastrin-releasing peptide receptor (GRPR), which is highly expressed in primary and metastasized prostate cancer. We have synthesized a new BN-based radiopharmaceutical 99mTechnetium-HYNIC(tricine/TPPTS)-Aca-BN(7–14) (99mTc-HABN) and evaluated its GRPR targeting properties in vitro and in a xenograft tumor model for human prostate cancer in athymic mice. Methods: 99mTc- HABN was synthesized and its lipophilicity and stability were investigated. The IC50, internalization and efflux properties were determined in vitro using the GRPR expressing human prostate cancer cell line PC-3. 99mTc-HABN biodistribution and microSPECT imaging were performed in PC-3 tumor-bearing athymic mice. Results: 99mTc-HABN was prepared with high labeling yield (>90%), high radiochemical purity (>95%) and a specific activity of ∼19.8 MBq/nmol. The partition coefficient log P value was −1.60±0.06. 99mTc-HABN proved to be stable in human serum for 6 hours. The IC50 of HABN was 12.81±0.14 nM. Incubation of PC-3 cells with 99mTc-HABN demonstrated rapid cellular internalization and a long intracellular retention time. When mice were injected with 99mTc-HABN the activity was predominantly cleared via the kidneys. Uptake in the tumor was 2.24±0.64 %ID/g after 30 minutes, with a steady decrease during the 4 hours study period. In vivo experiments with a blocking agent showed GRPR mediated uptake. 99mTc-HABN microSPECT imaging resulted in clear delineation of the tumor. Conclusions: 99mTc-HABN is a novel BN-based radiopharmaceutical that appears to be suitable for targeted imaging of prostate cancer. No significant financial relationships to disclose.

Effect of PEDF on the in vivo antitumor activities of low-dose chemotherapy in CRPC.

2013 ◽  
Vol 31 (6_suppl) ◽  
pp. 173-173
Author(s):  
Thomas Nelius ◽  
Everardo Cobos ◽  
Jennifer Hirsch ◽  
Stephanie Filleur
Keyword(s):  
Prostate Cancer ◽  
Low Dose ◽  
Pigment Epithelium ◽  
Growth Curves ◽  
Great Promise ◽  
Tumor Bearing ◽  
Cytotoxicity Effects ◽  
Low Dose Chemotherapy

173 Background: The development of metronomic/low dose administration of conventional chemotherapeutic drugs has shown great promise in the treatment of castration-refractory prostate cancer (CPRC). Pigment Epithelium-Derived Factor (PEDF) is a natural angio-inhibitor which is down-regulated in prostate cancer. We have previously demonstrated that the over-expression of PEDF in human CRPC PC3 cells decreased tumor growth in vivo. In the present study, we further validated PEDF anti-tumor properties in the highly metastatic CRPC LNCaP-derivative CL1 cells. We also hypothesized that PEDF may enhance the cytotoxicity effects of low dose docetaxel (DTX) and cyclophosphamide (CTX) chemotherapies in vivo. Methods: PC3 and CL1 cell lines were genetically modified to stably express the fluorescent DsRed Express protein with PEDF. Resulting cells were characterized in vitro for PEDF expression by western blot and, for proliferation by growth curves and clone formation in matrigel. PEDF anti-tumor effects were assessed on established s.c. xenografts in mice treated with DTX (5mg/kg ip every 4 days, 1mg/kg ip daily for 10 days, 0.5mg/kg every other day), CTX (10-20mg/kg in the drinking water) or placebo. Survival studies were performed by injecting CL1-PEDF or -control cells into the left lobe of the dorsal prostate of anesthetized mice. Results: We showed that PEDF expression inhibits the proliferation and induces the differentiation of CPRC cells in vitro, and decreases by 85% and 70% the development of s.c. PC3 and CL1 tumors, respectively. In the survival study, PEDF expression prolongs significantly (P=0.01; 95% confidence interval) the median survival of CL1 tumor-bearing mice (53±0.001 days versus 57±1). Furthermore, we demonstrated that PEDF enhances the cytotoxicity effects of low dose chemotherapy on established s.c. tumors (best Doc dose: 1mg/kg for PC3 and 5mg/kg for CL1; best CTX dose: 10mg/kg for PC3 and CL1) and prolongs significantly the survival of tumor-bearing mice undergoing low dose chemotherapy. Conclusions: These data reinforce the significance of PEDF as a potent target for the treatment of CRPC. It also emphasizes PEDF as a promising new agent to enhance the anti-tumor efficacy of low dose chemotherapies.

scholarly journals Improved pharmacokinetic and pharmacodynamic profile of a novel PEGylated native Erwinia chrysanthemi L-Asparaginase

2021 ◽  
Author(s):  
Tapasvi Modi ◽  
David Gervais
Keyword(s):  
Cell Lines ◽  
Half Life ◽  
Specific Activity ◽  
Kinetic Constant ◽  
Erwinia Chrysanthemi ◽  
Neutralising Antibodies ◽  
Pharmacodynamic Profile ◽  
Therapeutic Needs

SummaryIntroduction. Erwinase® (native Erwinia chrysanthemi L-Asparaginase (nErA)) is an approved second-line treatment for acute lymphoblastic leukaemia (ALL) in children and adolescents, who develop hypersensitivity or neutralising antibodies to E.coli derived L-Asparaginases (ASNases). However, nErA has a short in vivo half-life requiring frequent dosing schedules in patients. In this study, nErA was covalently conjugated to PEG molecules with the aim of extending its half-life in vivo. Methods. Firstly, efficacy of this novel product PEG-nErA was investigated on human ALL cell lines (Jurkat, CCRF-CEM and CCRF-HSB2), in vitro. Secondly, its pharmacokinetic (PK) and pharmacodynamic (PD) characteristics were determined, in vivo (12 rats in each group). Results. It was found that the specific activity (U/mg of enzyme) and the kinetic constant (KM) of nErA remained unaltered post PEGylation. PEG-nErA was shown to have similar cytotoxicity to nErA (IC50: 0.06–0.17 U/mL) on human ALL cell lines, in vitro. Further, when compared to nErA, PEG-nErA showed a significantly improved half-life in vivo, which meant that L-Asparagine (Asn) levels in plasma remained depleted for up to 25 days with a four-fold lower dose (100 U/kg) compared with 72 h for nErA at 400 U/kg dose. Conclusion. Overall, this next generation product PEG-nErA (with improved PK and PD characteristics compared to nErA) would bring a significant advantage to the therapeutic needs of ALL patients and should be further explored in clinical trials.

Evolution of Peptide-Based Prostate-Specific Membrane Antigen (PSMA) Inhibitors: An Approach to Novel Prostate Cancer Therapeutics

2020 ◽  
Vol 27 ◽  
Author(s):  
Andrew Siow ◽  
Renata Kowalczyk ◽  
Margaret A. Brimble ◽  
Paul W.R. Harris
Keyword(s):  
Prostate Cancer ◽  
Clinical Application ◽  
Rational Design ◽  
Therapeutic Potential ◽  
Rapid Development ◽  
Cancer Therapeutics ◽  
Prostate Specific Membrane Antigen ◽  
Specific Membrane

Background: Prostate cancer is one of the most common cancers worldwide, with approximately 1.1 million cases diagnosed annually. The rapid development of molecular imaging has facilitated greater structural understanding which can help formulate novel combination therapeutic regimens and more accurate diagnosis avoiding unnecessary prostate biopsies. This accumulated knowledge also provides greater understanding into aggressive stages of the disease and tumour recurrence. Recently, much progress has been made on developing peptidomimetic-based inhibitors as promising candidates to effectively bind to the prostate-specific membrane antigen (PSMA) which is expressed by prostate cancer cells. Objective: In this review, recent advances covering small-molecule and peptide-based PSMA inhibitors will be extensively reviewed providing a base for the rational design of future PSMA inhibitors. Method: Herein, we review the literature on selected PSMA inhibitors that have been developed from 1996-2020, emphasizing recent synthetic advances and chemical strategies whilst highlighting therapeutic potential and drawbacks of each inhibitor. Results: Synthesized inhibitors presented in this review demonstrate the clinical application of certain PSMA inhibitors exhibited in vitro and in vivo. Conclusion: This review highlights the clinical potential of PSMA inhibitors, analyzing the advantages and setbacks of the chemical synthetic methodologies utilized, setting precedence for the discovery of novel PSMA inhibitors for future clinical application.

scholarly journals On Site Production of [18F]-PSMA1007 Using Different [18F]-fluoride Activities. Practical, Technical and Economical Impact.

2021 ◽  
Author(s):  
Costantina Maisto ◽  
Anna Morisco ◽  
Roberta de Marino ◽  
Squame Elisabetta ◽  
Valentina Porfidia ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Large Scale ◽  
Radiochemical Purity ◽  
Radiochemical Yield ◽  
Good Target ◽  
Pet Ct ◽  
Fetal Bovine ◽  
Imaging Properties ◽  
Specific Membrane ◽  
Psma Inhibitor

Abstract BackgroundProstate-Specific Membrane Antigen is overexpressed in prostate cancer and it is considered a good target for staging of primary and recurrences as well as for radioligand therapy. Different PSMA-analogues have been investigated, labeled with [68Ga], showing excellent imaging properties; although, only small amounts can be produced for single synthesis. Recently, a fluorinated PSMA-inhibitor, [18F]-PSMA-1007, has been introduced, ensuring large-scale productions. In this study, the radiosynthesis of [18F]-PSMA-1007 using low (A), medium (B) and high (C) starting activities of [18F]-Fluoride, has been fully tested. The following parameters radiochemical yield, radiochemical purity and stability of [18F]-PSMA-1007 have been measured in 65, consecutive batches to verify the effects of the three different conditions. In addition, the analysis of the costs for the production has been performed. ResultsThe radiochemical yield percentage for low and medium range of activities of [18F]-Fluoride was 52%, while for the high range it decreases to 40%. The radiochemical purity was 99% in all three tested starting activities. [18F]-PSMA-1007 did not show radiolysis up to 8 hours after the end of synthesis, confirming that the radiopharmaceutical is stable an suitable for PET studies in humans. Furthermore, stability studies performed in fetal bovine serum demonstrated radiochemical stability at 37°C for 120’. ConclusionsA starting activity of [18F]-Fluoride of 90 GBq (range B) enables a final amount of [18F]-PSMA-1007 of about 50 GBq, which is powerful for different choices: to perform up to 25 PET/CT scans in a referral institution for prostate cancer, and/or to supply the eventual peripheral PET centers.

Evidence for the existence of a C25-sesterterpene pathway of steroidogenesis not involving cholesterol as an obligatory intermediate

1983 ◽  
Vol 61 (7) ◽  
pp. 714-721 ◽  
Author(s):  
Bhagu R. Bhavnani ◽  
Tony Wong
Keyword(s):  
Isotope Dilution ◽  
Guinea Pigs ◽  
Radiochemical Purity ◽  
Specific Activity ◽  
Dilution Technique ◽  
Isotope Dilution Technique ◽  
Alternate Pathway

Previous in vitro studies had indicated the possibility of steroidogenesis through a C25-sesterterpene pathway in which squalene and cholesterol are not required as obligatory intermediates. To investigate whether such a pathway exists in vivo, the precursor role of [7-3H]23,24-dinor-5-cholene-3β,20-diol in the in vivo formation of cortisol by the guinea pigs was studied. The [3H]23,24-dinor-5-cholene-3β,20-diol was synthesized by reacting [3H]pregnenolone acetate with a Grignard reagent. The product was purified by chromatography and its radiochemical purity was established by the isotope dilution technique. In the first experiment a total of 134 × 106 dpm of [3H]23,24-dinor-5-cholene-3β,20-diol was injected subcutaneously into three guinea pigs. Urine was collected for 8 days and was pooled. Only 12% of the administered dose was excreted in the urine. The urine was extracted and a neutral extract (3 × 106 dpm) was obtained. From this extract 2.3 mg of cortisol containing 2.9 × 104 dpm was isolated. Radiochemical purity of the isolated cortisol was established by the isotope dilution technique. Radiochemical purity was further confirmed by conversion to cortisol 21-acetate and subsequently to 11β-hydroxyandrost-4-ene-3,17-dione and recrystallization to constant specific activity. The results of this experiment were confirmed by repeating the experiment with a higher specific activity [3H]23,24-dinor-5-cholene-3β,20-diol. These results indicate that the C25-sesterterpene pathway is a possible in vivo alternate pathway of steroidogenesis, not involving either squalene or cholesterol as obligatory intermediates.

scholarly journals Preparation and Biological Evaluation of [99mTc]Tc-CNGU as a PSMA-Targeted Radiotracer for the Imaging of Prostate Cancer

Molecules ◽  
2020 ◽  
Vol 25 (23) ◽  
pp. 5548
Author(s):  
Di Xiao ◽  
Xiaojiang Duan ◽  
Qianqian Gan ◽  
Xuran Zhang ◽  
Junbo Zhang
Keyword(s):  
Prostate Cancer ◽  
Biological Evaluation ◽  
Log P ◽  
Tumor Uptake ◽  
Biodistribution Studies ◽  
Biological Target ◽  
High Tumor Uptake ◽  
Specific Membrane ◽  
Psma Inhibitor

Prostate-specific membrane antigen (PSMA) is a well-established biological target that is overexpressed on the surface of prostate cancer lesions. Radionuclide-labeled small-molecule PSMA inhibitors have been shown to be promising PSMA-specific agents for the diagnosis and therapy of prostate cancer. In this study, a glutamate-urea-based PSMA-targeted ligand containing an isonitrile (CNGU) was synthesized and labeled with 99mTc to prepare [99mTc]Tc-CNGU with a high radiochemical purity (RCP). The CNGU ligand showed a high affinity toward PSMA (Ki value is 8.79 nM) in LNCaP cells. The [99mTc]Tc-CNGU exhibited a good stability in vitro and hydrophilicity (log P = −1.97 ± 0.03). In biodistribution studies, BALB/c nude mice bearing LNCaP xenografts showed that the complex had a high tumor uptake with 4.86 ± 1.19% ID/g, which decreased to 1.74 ± 0.90% ID/g after a pre-injection of the selective PSMA inhibitor ZJ-43, suggesting that it was a PSMA-specific agent. Micro-SPECT imaging demonstrated that the [99mTc]Tc-CNGU had a tumor uptake and that the uptake was reduced in the image after blocking with ZJ-43, further confirming its PSMA specificity. All of the results in this work indicated that [99mTc]Tc-CNGU is a promising PSMA-specific tracer for the imaging of prostate cancer.

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